An essential aspect of this PPG is to maintain a state-of-the-art core facility that supports the imaging needs of the four proposed projects. Similar to the work performed in the initial funding cycle, we propose to make use of a variety of microscopic and macroscopic imaging technologies to gain a better understanding of biofilm assembly and metabolism, and how these affect the host immune response both in vitro and in vivo. Thus, the primary function of this Bioimaging Core will be to maintain the cutting-edge instrumentation needed to visualize the specific transcriptional and metabolic changes that occur during biofilm development, and to assess the impact of these changes in the host environment. The first specific aim of this core is to upgrade and maintain the Center for Staphylococcal Research (CSR) confocal microscopy facility. This aim will provide an upgrade for the existing Zeiss LSM510 META confocal microscope utilized by all members of our group to the more powerful LSM710 series, thus, providing optimal imaging capabilities. The second specific aim will be to provide the maintenance and expertise needed to analyze biofilm development using our BioFlux Microfluidics System. This aim will be primarily utilized by Projects 1 and 3 to assess the effects of various metabolic and regulatory mutations on biofilm development and gene expression. The third and final specific aim will be to support our In Vivo Imaging System (IVIS) Spectrum instrument, which will be used by Projects 2, 3, and 4 to visualize the progression of infection in live animals. All three of these aims will provide essential technology needed to successfully execute the goals of the projects described in this proposal.